The goals of the research are to understand aspects of phospholipid and glycolipid metabolism that are relevant for the development of megakaryocytes and the production of platelets. Our studies indicate that megakaryocytes have a greater capacity for the metabolism of unsaturated fatty acids than platelets and that uptake of unsaturated fatty acids occurs primarily in younger megakaryocytes. The proposed research will extend these studies and determine the relative roles of megakaryocytes and platelets for the establishment of the highly specific arrangement of fatty acids that exists in both cells. These studies will be performed in isolated guinea pig megakaryocytes and in subpopulations of megakaryocytes enriched with younger and older cells, prepared by density gradient centrifugation, to relate our findings to megakaryocyte maturity. The metabolism of the following unsaturated fatty acids will be studied: oleic, linoleic, arachidonic and eicosapentaenoic acids. To define fatty acid metabolism and mechanisms for the regulation of fatty acids in megakaryocytes, the following will be investigated: 1) Synthesis, uptake and transfer of unsaturated fatty acids amongst megakaryocyte phospholipids; 2) the distribution of newly-synthesized and incorporated fatty acids in megakaryocyte phospholipids in comparison to that in platelets; 3) the regulation of synthesis, uptake and transfer by saturated and unsaturated fatty acids. To determine the enzymatic basis for fatty acid uptake and transfer and the regulation of these events, acyltransferase and transacylase activities will be studied in megakaryocytes. We also have studied the exposure of glycolipids and gangliosides on the surface of platelets. Preliminary data indicate that it is feasible to study megakaryocyte glycolipids. The proposed research will characterize glycolipids, sulfatides and gangliosides in megakaryocytes and also will identify these lipids in megakaryocytes at different stages of maturation. The expression of gangliosides and neutral glycolipids on the surface of megakaryocytes at various stages of development will be defined by external labeling techniques. Glycolipids are known to be important for cellular maturation but as yet there is no information about these unique lipids in megakaryocytes. We will characterize the two sulfatides that we have detected in human platelets in respect to sphingosine, fatty acid and carbohydrate composition, determine whether they are components of platelet cytoskeletons, and study their ability to provide a surface for the initiation of the intrinsic coagulation system.